Article

Abnormal surface liquid pH regulation by cultured cystic fibrosis bronchial epithelium.

Cystic Fibrosis/Pulmonary Research and Treatment Center, School of Medicine, University of North Carolina, Chapel Hill, NC 27599-7248, USA.
Proceedings of the National Academy of Sciences (Impact Factor: 9.81). 01/2004; 100(26):16083-8. DOI: 10.1073/pnas.2634339100
Source: PubMed

ABSTRACT Cystic fibrosis (CF) transmembrane conductance regulator (CFTR)-dependent airway epithelial bicarbonate transport is hypothesized to participate in airway surface liquid pH regulation and contribute to lung defense. We measured pH and ionic composition in apical surface liquid (ASL) on polarized normal (NL) and CF primary bronchial epithelial cell cultures under basal conditions, after cAMP stimulation, and after challenge with luminal acid loads. Under basal conditions, CF epithelia acidified ASL more rapidly than NL epithelia. Two ASL pH regulatory paths that contributed to basal pH were identified in the apical membrane of airway epithelia, and their activities were measured. We detected a ouabain-sensitive (nongastric) H+,K+-ATPase that acidified ASL, but its activity was not different in NL and CF cultures. We also detected the following evidence for a CFTR-dependent HCO3- secretory pathway that was defective in CF: (i). ASL [HCO3-] was higher in NL than CF ASL; (ii). activating CFTR with forskolin/3-isobutyl-1-methylxanthine alkalinized NL ASL but acidified CF ASL; and (iii). NL airway epithelia more rapidly and effectively alkalinized ASL in response to a luminal acid challenge than CF epithelia. We conclude that cultured human CF bronchial epithelial pHASL is abnormally regulated under basal conditions because of absent CFTR-dependent HCO3- secretion and that this defect can lead to an impaired capacity to respond to airway conditions associated with acidification of ASL.

0 Bookmarks
 · 
110 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Mucins are essential components in mucus gels that form protective barriers at all epithelial surfaces, but much remains unknown about their assembly, intragranular organization and post-secretion unfurling to form mucus. MUC5B is a major polymeric mucin expressed by respiratory epithelia and we investigated the molecular mechanisms involved during its assembly. Studies of intact polymeric MUC5B revealed a single high affinity calcium-binding site, distinct from multiple low affinity sites on each MUC5B monomer. Self-diffusion studies with intact MUC5B showed that calcium binding at the protein-site catalyzed reversible cross-links between MUC5B chains to form networks. The site of cross-linking was identified in the MUC5B D3 domain as it was specifically blocked by D3 peptide antibodies. Biophysical analysis and single particle EM of recombinant MUC5B N-terminus (D1D2D'D3; NT5B) and sub-domains (D1, D1-D2, D2-D'-D3 and D3) generated structural models of monomers and disulfide-linked dimers and suggested that MUC5B multimerizes by disulfide-linkage between D3 domains to form linear polymer chains. Moreover, these analyses revealed reversible homotypic interactions of NT5B at low pH and in high calcium, between disulfide-linked NT5B dimers, but not monomers. These results enable a model of MUC5B to be derived, which predicts mechanisms of mucin intracellular assembly and storage, which may be common to the other major gel forming polymeric mucins.
    Journal of Biological Chemistry 04/2014; · 4.65 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Disrupted HCO3(-) transport and reduced airway surface liquid (ASL) pH in cystic fibrosis (CF) may initiate airway disease. We hypothesized that ASL pH is reduced in neonates with CF. In neonates with and without CF, we measured pH of nasal ASL. We also measured nasal pH in older children and adults. In neonates with CF, nasal ASL (pH5.2±0.3) was more acidic than in non-CF neonates (pH6.4±0.2). In contrast, nasal pH of CF children and adults was similar to values measured in people without CF. At an age when infection, inflammation and airway wall remodeling are minimal, neonates with CF had an acidic nasal ASL compared to babies without CF. The CF:non-CF pH difference disappeared in older individuals, perhaps because secondary manifestations of disease increase ASL pH. These results aid understanding of CF pathogenesis and suggest opportunities for therapeutic intervention and monitoring of disease.
    Journal of cystic fibrosis: official journal of the European Cystic Fibrosis Society 01/2014; · 3.19 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: The epithelia that line the conducting airways are the lung's first point of contact with inhaled pathogens and toxicants. As such, they are known to play an important role in the lung's innate defense system, which includes (i) the production of airway surface liquid (ASL) that helps cleanse the airways through the physical removal of pathogens and toxicants on the mucociliary escalator and (ii) the secretion of anti-microbial proteins into the ASL to kill inhaled pathogens. Interestingly, the recently crystalized short palate lung and nasal epithelial clone 1 (SPLUNC1) protein appears to be a multi-functional protein. That is, it not only acts as an anti-microbial agent, but also modulates ASL homeostasis by acting as an endogenous inhibitor of the epithelial Na(+) channel (ENaC). This review will focus on the latter function of SPLUNC1, and will discuss new structural and physiological data regarding SPLUNC1's failure to function as a regulator of ASL hydration in CF airways.
    The international journal of biochemistry & cell biology 03/2014; · 4.89 Impact Factor

Full-text

View
0 Downloads
Available from